Proteases
In general, proteases typically affect protein cleavage, processing, and/or turnover. Proteases may be categorized into families by the different amino acid sequences (generally between 2 and 10 residues) located on either side of the cleavage site of the protease.
The proper functioning of the cell requires careful control of the levels of important structural proteins, enzymes, and regulatory proteins. One of the ways that cells can reduce the steady state level of a particular protein is by proteolytic degradation. Further, one of the ways cells produce functioning proteins is to produce pre- or pro-protein precursors that are processed by proteolytic degradation to produce an active moiety. Thus, complex and highly regulated mechanisms have been evolved to accomplish this degradation.
Proteases regulate many different cell proliferation, differentiation, and signaling processes by regulating protein turnover and processing. Uncontrolled protease activity (either increased or decreased) has been implicated in a variety of disease conditions including inflammation, cancer, arteriosclerosis, and degenerative disorders.
An additional role of intracellular proteolysis is in the stress-response. Cells that are subject to stress such as starvation, heat-shock, chemical insult or mutation respond by increasing the rates of proteolysis. One function of this enhanced proteolysis is to salvage amino acids from non-essential proteins. These amino acids can then be re-utilized in the synthesis of essential proteins or metabolized directly to provide energy. Another function is in the repair of damage caused by the stress. For example, oxidative stress has been shown to damage a variety of proteins and cause them to be rapidly degraded.
The International Union of Biochemistry and Molecular Biology (IUBMB) has recommended to use the term peptidase for the subset of peptide bond hydrolases (Subclass E.C 3.4.). The widely used term protease is synonymous with peptidase. Peptidases comprise two groups of enzymes: the endopeptidases and the exopeptidases, which cleave peptide bonds at points within the protein and remove amino acids sequentially from either N or C-terminus respectively. The term proteinase is also used as a synonym word for endopeptidase and four mechanistic classes of proteinases are recognized by the IUBMB: two of these are described below (also see: Handbook of Proteolytic Enzymes by Barrett, Rawlings, and Woessner AP Press, NY 1998). Also, for a review of the various uses of proteases as drug targets, see: Weber M, Emerging treatments for hypertension: potential role for vasopeptidase inhibition; Am J Hypertens 1999 November;12(11 Pt 2):139S–147S; Kentsch M, Otter W, Novel neurohormonal modulators in cardiovascular disorders. The therapeutic potential of endopeptidase inhibitors, Drugs R D 1999 April;1(4):331–8; Scarborough R M, Coagulation factor Xa: the prothrombinase complex as an emerging therapeutic target for small molecule inhibitors, J Enzym Inhib 1998;14(1):15–25; Skotnicki J S, et al., Design and synthetic considerations of matrix metalloproteinase inhibitors, Ann N Y Acad Sci 1999 Jun. 30;878:61–72; McKerrow J H, Engel J C, Caffrey C R, Cysteine protease inhibitors as chemotherapy for parasitic infections, Bioorg Med Chem 1999 April;7(4):639–44; Rice K D, Tanaka R D, Katz B A, Numerof R P, Moore W R, Inhibitors of tryptase for the treatment of mast cell-mediated diseases, Curr Pharm Des 1998 October;4(5):381–96; Materson B J, Will angiotensin converting enzyme genotype, receptor mutation identification, and other miracles of molecular biology permit reduction of NNT Am J Hypertens 1998 August;11(8 Pt 2):138S–142S.
Cathepsin S
Cathepsin S is a lysosomal cysteine protease that catalyzes the removal of the invariant chain (Ii) from MHC class II molecules (Morton, P. A., Zacheis, M. L., Giacoletto, K. S., Manning, J. A., Schwartz, B. D., (1995) J. Immunol. 154, 137–150). Ii serves dual functions. Ii serves as a molecular chaperon that promotes the MHC class II/Ii complex exit from the ER to the endosomal system through the secretory pathway. Ii also functions as a molecular inhibitor that occupies the peptide-binding pocket of MHC class II to prevent premature binding of native peptides in the secretory pathway (Hsieh, C. S., deRoos, P., Honey, K., Beers, C., and Rudensky, A. Y. (2002) J. Immunol. 168, 2618–2625). Experiments using protease inhibitors and knockout mice have clearly shown that cathepsin S is involved in proteolytic clearance of Ii from MHC class II molecules (Riese, R. J., Mitchell, R. N., Villadangos, J. A., Shi, G. P., Palmer, J. T., Karp, E. R., De Sanctis, G. T., Ploegh, H. L., and Chapman, H. A. (1998) J. Clin. Invest. 101, 2351–2363; Nakagawa, T. Y., Brissette, E. H., Lira, P. D., Griffiths, R. J., Petrushova, N., Stock, J., McNeish, J. D., Eastman, S. E., Howard, E. D., Clarke, S. R., Rosloniec, E. F., Elliott, E. A., and Rudensky, A. Y. (1999) Immunity 10, 207–217). Inhibition of cathepsin S precludes antigen loading to MHC class II molecules and disables the antigen-presenting cell, thereby preventing the antigen-presenting cell from presenting antigen to CD4+ T cells (Shi, G.-P., Villadangos, J. A., Dranoff, G., Small, C., Gu, L., Haley, K. J., Rises, R., Ploegh, H. L., Chapman, H. A. (1999) Immunity 10, 197–206). Selective inhibition of cathepsin S is therefore considered to be therapeutically useful to attenuate the elevated immune responses found in many autoimmune disorders.
To investigate the effectiveness of cathepsin S inhibitors, it is typically necessary to evaluate species-specific cathepsin S activity in different animal autoimmune disease models. In particular, to develop cathepsin S inhibitors for the treatment of human disorders, it is typically desirable to test candidate inhibitor compounds against cathepsin S proteins of non-human primates, such as monkeys. Non-human primates, because they are more closely related to humans than rodents and other laboratory animals, can more accurately predict the effectiveness (as well as toxicity and other undesirable side effects) of the inhibitor compound in humans, and are therefore advantageous for use as animal models for evaluating the use of potential therapeutic compounds for treating human disease, prior to administration of the compounds to humans.
Consequently, because cathepsin S proteins are well established in the art as playing key roles in important human diseases (such as autoimmune disorders), and because monkeys serve as an optimal animal model for evaluating candidate therapeutic compounds for the treatment of human diseases, a need exists in the art for isolated cathepsin S proteins, and encoding nucleic acid molecules, from non-human primates such as monkeys.
Proteases and Cancer
Proteases are critical elements at several stages in the progression of metastatic cancer. In this process, the proteolytic degradation of structural protein in the basal membrane allows for expansion of a tumor in the primary site, evasion from this site as well as homing and invasion in distant, secondary sites. Also, tumor induced angiogenesis is required for tumor growth and is dependent on proteolytic tissue remodeling. Transfection experiments with various types of proteases have shown that the matrix metalloproteases play a dominant role in these processes in particular gelatinases A and B (MMP-2 and MMP-9, respectively). For an overview of this field, see Mullins, et al., Biochim. Biophys. Acta 695, 177, 1983; Ray, et al., Eur. Respir. J. 7, 2062, 1994; Birkedal-Hansen, et al., Crit. Rev. Oral Biol. Med. 4, 197, 1993.
Furthermore, it was demonstrated that inhibition of degradation of extracellular matrix by the native matrix metalloprotease inhibitor TIMP-2 (a protein) arrests cancer growth (DeClerck, et al., Cancer Res. 52, 701, 1992) and that TIMP-2 inhibits tumor-induced angiogenesis in experimental systems (Moses, et al. Science 248, 1408, 1990). For a review, see DeClerck, et al., Ann. N. Y. Acad. Sci. 732, 222, 1994. It was further demonstrated that the synthetic matrix metalloprotease inhibitor batimastat when given intraperitoneally inhibits human colon tumor growth and spread in an orthotopic model in nude mice (Wang, et al. Cancer Res. 54, 4726, 1994) and prolongs the survival of mice bearing human ovarian carcinoma xenografts (Davies, et. al., Cancer Res. 53, 2087, 1993). The use of this and related compounds has been described in Brown, et al., WO-9321942 A2.
There are several patents and patent applications claiming the use of metalloproteinase inhibitors for the retardation of metastatic cancer, promoting tumor regression, inhibiting cancer cell proliferation, slowing or preventing cartilage loss associated with osteoarthritis or for treatment of other diseases as noted above (e.g. Levy, et al., WO-9519965 A1; Beckett, et al., WO-9519956 A1; Beckett, et al., WO-9519957 A1; Beckett, et al., WO-9519961 A1; Brown, et al., WO-9321942 A2; Crimmin, et al., WO-9421625 A1; Dickens, et al., U.S. Pat. No. 4,599,361; Hughes, et al., U.S. Pat. No. 5,190,937; Broadhurst, et al., EP 574758 A1; Broadhurst, et al., EP 276436; and Myers, et al., EP 520573 A1.
Mammalian Models of Human Disease
Non-human primates such as monkeys, as well as rodents such as mice (Mus musculus) and other animals such as rabbits and guinea pigs, are commonly used in biomedical research as model systems for studying human diseases and for developing therapeutic and diagnostic agents for human diseases. Detailed descriptions of techniques and protocols for manipulating and using such animal models of human disease, particularly mouse models, such as techniques for using homologous recombination to produce mutant mouse strains and mutant cell lines with specific genes inactivated (e.g., “knockout” mice), are readily available, and the techniques described for mice can be applied to monkeys and other non-human primates as well as other laboratory animals. For example, for a review of mouse models for studying gene/protein functions and interactions, cell biology, and human diseases, including mouse mutagenesis techniques currently used in the art, see Current Protocols in Molecular Biology, John Wiley & Sons, Inc., “Manipulating the Mouse Genome”, chapter 23, supplements 51–53 (2000–2001); Nolan et al., Nat Genet 2000 August;25(4):440–3; Brown, J Inherit Metab Dis 1998 August;21(5):532–9; Nolan, Pharmacogenomics 2000 August; 1(3):243–55; Justice, Nat Rev Genet 2000 November; 1(2): 109–15; and Mansuy et al., Exp Physiol 2000 November;85(6):661–79.
Protease proteins, particularly cathepsin S proteases, are a major target for drug action and development, and the monkey provides an optimal animal model in which to study these protease proteins and the diseases with which they are associated. Accordingly, it is valuable to the field of pharmaceutical development to identify and characterize previously unknown members of this subfamily of protease proteins in the monkey. The present invention advances the state of the art by providing isolated monkey cathepsin S proteins and encoding nucleic acid molecules.